TRPML1 gating modulation by allosteric mutations and lipids (Design of allosteric mutations that recapitulate the gating of TRPML1)
Curation statements for this article:-
Curated by eLife
eLife assessment
Transient receptor potential mucolipin 1 (TRPML1) functions as a lysosomal ion channel whose variants are associated with lysosomal storage disorder mucolipidosis type IV. This important report describes local and global structural changes driven by the binding of regulatory phospholipids and by mutations allosteric that allosterically cause gain or loss of channel function. Most of the claims related to the allosteric regulation of TRPML1 have solid support by two new cryo-EM structures, that of the gain of function Y404W mutant and that of the wild-type channel bound to the inhibitor PI(4,5)P2. The new cryo-EM findings are evaluated within the context of previously reported TRPML1 structures, and a proposed allosteric gating mechanism is partially supported by functional electrophysiology results.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Reading list (BiophysicsColab)
Abstract
Transient Receptor Potential Mucolipin 1 (TRPML1) is a lysosomal cation channel whose loss-of-function mutations directly cause the lysosomal storage disorder mucolipidosis type IV (MLIV). TRPML1 can be allosterically regulated by various ligands including natural lipids and small synthetic molecules and the channel undergoes a global movement propagated from ligand-induced local conformational changes upon activation. In this study, we identified a functionally critical residue, Tyr404, at the C-terminus of the S4 helix, whose mutations to tryptophan and alanine yield gain- and loss-of-function channels, respectively. These allosteric mutations mimic the ligand activation or inhibition of the TRPML1 channel without interfering with ligand binding and both mutant channels are susceptible to agonist or antagonist modulation, making them better targets for screening potent TRPML1 activators and inhibitors. We also determined the high-resolution structure of TRPML1 in complex with the PI(4,5)P 2 inhibitor, revealing the structural basis underlying this lipid inhibition. In addition, an endogenous phospholipid likely from sphingomyelin is identified in the PI(4,5)P 2 -bound TRPML1 structure at the same hotspot for agonists and antagonists, providing a plausible structural explanation for the inhibitory effect of sphingomyelin on agonist activation.
Article activity feed
-
-
-
eLife assessment
Transient receptor potential mucolipin 1 (TRPML1) functions as a lysosomal ion channel whose variants are associated with lysosomal storage disorder mucolipidosis type IV. This important report describes local and global structural changes driven by the binding of regulatory phospholipids and by mutations allosteric that allosterically cause gain or loss of channel function. Most of the claims related to the allosteric regulation of TRPML1 have solid support by two new cryo-EM structures, that of the gain of function Y404W mutant and that of the wild-type channel bound to the inhibitor PI(4,5)P2. The new cryo-EM findings are evaluated within the context of previously reported TRPML1 structures, and a proposed allosteric gating mechanism is partially supported by functional electrophysiology results.
-
Reviewer #1 (Public Review):
In their manuscript, Gan and colleagues identified a functional critical residue, Tyr404, which when mutated to W or A results in GOF and LOF of TRPML1 activity, respectively. In addition, the authors provide a high-resolution structure of TRPML1 with PI(4,5)P2 inhibitor. This high-resolution structure also revealed a bound phospholipid likely sphingomyelin at the agonist/antagonist site, providing a plausible explanation for sphingomyelin inhibition of TRPML1.
This is an interesting study, revealing valuable additional information on TRPML1 gating mechanisms including effects on endogenous phospholipids on channel activity. The provided data are convincing. Some major open questions remain. The work will be of interest to a wide audience including industry researchers occupied with TRPML1 exploration as a …
Reviewer #1 (Public Review):
In their manuscript, Gan and colleagues identified a functional critical residue, Tyr404, which when mutated to W or A results in GOF and LOF of TRPML1 activity, respectively. In addition, the authors provide a high-resolution structure of TRPML1 with PI(4,5)P2 inhibitor. This high-resolution structure also revealed a bound phospholipid likely sphingomyelin at the agonist/antagonist site, providing a plausible explanation for sphingomyelin inhibition of TRPML1.
This is an interesting study, revealing valuable additional information on TRPML1 gating mechanisms including effects on endogenous phospholipids on channel activity. The provided data are convincing. Some major open questions remain. The work will be of interest to a wide audience including industry researchers occupied with TRPML1 exploration as a drug target.
-
Reviewer #2 (Public Review):
The transient receptor potential mucolipin 1 (TRPML1) functions as a lysosomal organelle ion channel whose variants are associated with lysosomal storage disorder mucolipidosis type IV. Understanding sites that allosterically control the TRPML1 channel function may provide new molecular moieties to target with prototypic drugs.
Gan et al provide the first high-resolution cryo-EM structures of the TRPML1 channel (Y404W) in the open state without any activating ligands. This new structure demonstrates how a mutation at a site some distance away from the pore can influence the channel's conducting state. However, the authors do not provide a structural analysis of the Y404W pore which would validate their open-state claims. Nonetheless, Gan et al provide compelling electrophysiology evidence which supports the …
Reviewer #2 (Public Review):
The transient receptor potential mucolipin 1 (TRPML1) functions as a lysosomal organelle ion channel whose variants are associated with lysosomal storage disorder mucolipidosis type IV. Understanding sites that allosterically control the TRPML1 channel function may provide new molecular moieties to target with prototypic drugs.
Gan et al provide the first high-resolution cryo-EM structures of the TRPML1 channel (Y404W) in the open state without any activating ligands. This new structure demonstrates how a mutation at a site some distance away from the pore can influence the channel's conducting state. However, the authors do not provide a structural analysis of the Y404W pore which would validate their open-state claims. Nonetheless, Gan et al provide compelling electrophysiology evidence which supports the proposed Y404W gain of function effect. The authors propose an allosteric mechanism with the following molecular details- the Y404 to W sidechain substitution provides extra van der Waals contacts within the pocket surrounded by helices of the VSD-like domain and causes S4 bending which in turn opens to the pore through the S4-S5 linker. Conversely, the author functionally demonstrates that an alanine mutation at this site causes a loss of function. Although the authors do not provide a structure of the Y404A mutation, they propose that the alanine substitution disrupts the sidechain packing and likely destabilizes the open conformation. TRPM1 channels are regulated by PIP2 species, which is related to their cell function. In the membrane of lysosomes, PI(3,5)P2 activates the channel, whereas PI(4,5)P2 found in the plasma membrane has inhibitory effects. To understand its lipid regulation, the authors solved a cryo-EM structure of TRPM1 bound to PI(4,5)P2 in its presumed closed state. Again, while the provided functional evidence suggests that PI(4,5)P2 occupancy inhibits TRPML1 current, the authors do not provide analysis of the pore which would support their closed state assertion. Within this same structure, the authors observe a density that may be attributed to sphingomyelin (or possibly phosphocholine). Using electrophysiology of WT and the Y404W channels, the authors report sphingomyelins antagonist effect on TRPML1 currents under low luminal (external) pH. Taken together, the results described in Gan et al provide compelling evidence for a gating (open, closed) mechanism of the TRPML1 pore which can be allosterically regulated by altered packing and lipid interactions within the VSDL.
-
